Biodiesel, monoalkyl esters of long-chain fatty acids with short-chain alcohols derived from triacylglycerols (TAGs), can be produced from renewable biomass sources (Canakci and Sanli, J Ind Microbial Biotechnol 2008, 35:431-441, Du et al., Appl Microbiol Biotechnol 2008, 79:331-337, Vasudevan and Briggs, J Ind Microbial Biotechnol 2008, 35:421-430). Reports on the production of microbial oils by oleaginous microorganisms such as yeast, fungi, bacteria and microalgae have increased due to heightened awareness of the needs for alternative oil sources in the form of high energy molecules from renewable resources (Alvarez and Steinbüchel, Appl Microbiol Biotechnol 2002, 60:367-376, Antoni et al., Appl Microbiol Biotechnol 2007, 77:23-35; Li et al., Appl Microbiol Biotechnol 2008, 80:749-756; Hu et al., Plant J 2008, 54:621-639).
Rhodococcus opacus PD630 (R. opacus PD630) grown on gluconate medium is capable of accumulating TAGs up to 76% of the cell dry weight (CDW) (Alvarez et al., Arch Microbiol 1996, 165:377-386, Wältermann et al, Microbiology 2000, 146:1143-1149). It has also been reported that R. opacus PD630 grown in a fed-batch condition on a medium containing sugar beet molasses and sucrose as carbon sources reached a cell density of 37.4 g CDW l−1 with a fatty acid content of 51.9% of the CDW (Voss and Steinbüchel, Appl Microbiol Biotechnol 2001, 55:547-555).
Currently, the development of second generation biofuel technologies that can be produced sustainably by using lignocellulosic biomass have been accelerated due to food-fuel conflict avoidance and use of wastes or unutilized materials (Stein, J Am Diet Assoc 2007, 107:1870-1878; Tollefson, Nature 2008, 451:880-883). Lignocellulosic biomass includes xylan that is a xylose polymer. However, the use of xylose-based media for the growth of R. opacus PD630 cultures has not been possible, owing to the inability of R. opacus PD630 to metabolize xylose.
Similarly, synthesis of biodiesel from plant oil has led to the production of large quantities of glycerol. As world production of biodiesel is increasing exponentially, large quantities of low cost glycerol that could be used as a substrate for bioprocesses will be available. Although R. opacus PD630 can utilize glucose and other carbon sources for growth, it does not utilize glycerol as sole carbon source.